- Email: [email protected]
Synthesis and antiepileptic studies of new imidazo[1,2-a]pyridine derivatives
G Model
CCLET-2620; No. of Pages 4 Chinese Chemical Letters xxx (2013) xxx–xxx
Contents lists available at SciVerse ScienceDirect
Chinese Chemical Letters journal homepage: www.elsevier.com/locate/cclet
Original article
Synthesis and antiepileptic studies of new imidazo[1,2-a]pyridine derivatives Shrikanth Ulloora a, Airody Vasudeva Adhikari a,*, Ramakrishna Shabaraya b a b
Department of Chemistry, National Institute of Technology Karnataka, Surathkal, Mangalore 575025, Karnataka, India Srinivas College of Pharmacy, Valachil, Mangalore, Karnataka, India
A R T I C L E I N F O
A B S T R A C T
Article history: Received 7 March 2013 Received in revised form 6 May 2013 Accepted 17 May 2013 Available online xxx
In this manuscript, we report the synthesis of newly designed imidazo[1,2-a]pyridines carrying active pharmacophores as potential anticonvulsant agents. Newly synthesized target compounds were characterized by FTIR, 1H NMR, 13C NMR, and mass spectroscopy followed by elemental analysis studies. Preliminary anticonvulsant screening study of target compounds was carried out following MES and scPTZ methods. Compounds 6e and 6f, possessing electron rich aryl substituent at position-2 and tolyl substituted oxazolone moiety at the position-3 of imidazo[1,2-a]pyridine ring, exhibited activity comparable to standard drug diazepam and emerged as lead compounds. New compounds displayed enhanced activity in scPTZ method, indicating their ability to raise the seizure threshold. Their neurotoxicity study by Rotarod test showed that they are nontoxic at all tested doses. ß 2013 Airody Vasudeva Adhikari. Published by Elsevier B.V. on behalf of Chinese Chemical Society. All rights reserved.
Keywords: Anticonvulsant study Imidazo[1,2-a]pyridine In vivo study Neurotoxicity
1. Introduction Epilepsy is a major neurological disease, affecting about 50 million people worldwide [1]. The cellular mechanism of human epilepsy is still uncertain and hence present drug therapy only treats epileptic symptoms, rather than curing it [2]. Now there are more than 40 different anti-epileptic drugs (AEDs) in clinical use, but still about 30% of patients continue to experience uncontrolled seizures, and they are pharmaco-resistant to the available therapy [3]. Further, present medication for seizures requires continuous medication for a long period, which is in turn associated with many adverse side effects, such as nausea, ataxia, drowsiness, gastrointestinal disturbance, hyperplasia, anaemia etc. [4]. As a result, design and development of new, efficient antiepileptic agents has become an active area of research in medicinal chemistry. Imidazo[1,2-a]pyridines are an important class of non-benzodiazepines, exhibiting very good CNS activities and showing fewer adverse effects compared to classical benzodiazepines [5]. Some of the imidazo[1,2-a]pyridine derivatives, such as Zolpidem, Alpidem, Saripidem and DS-1, are well-known CNS drugs, exhibiting antiepileptic activity at lesser doses [6]. The pharmacological profiles of imidazo[1,2-a]pyridines are mainly dependent on the nature of the substituents at their position-2 and position-3 [7]. The imidazo[1,2-a]pyridines carrying aryl groups at the position-2 are reported as highly CNS active
* Corresponding author. E-mail address: [email protected] (A.V. Adhikari).
scaffolds [8]. Further, it is also revealed that, such scaffolds possessing a hydrophobic unit at the position-8 along with a halogen substituent on the aryl ring can bind selectively to benzodiazepine receptors [9]. As such, this work presents new imidazo[1,2-a]pyridines carrying a halo substituted aryl ring at the position-2 along with a hydrophobic methyl group at the position8. Additionally, active pharmacophoric groups like oxazolones, pyrazolones, imines and hydroxyl derivatives have been incorporated at the position-3 owing to their established prominent anticonvulsant activities [10,11]. Accordingly, four new series of imidazo[1,2-a]pyridines carrying imines (3a–d), enol derivatives (5a–d), oxazolones (6a–f), carboxylate intermediates (7a–c) and pyrazolones (8a–c) have been synthesized following appropriate synthetic routes as shown in Scheme 1. 2. Experimental The synthesis and the characterization details of intermediates (1a–c and 2a–c) were reported in our previous publication [12]. The imines 3a–d were obtained by condensing aldehydes 2a–c with appropriate aromatic amines in the presence of a catalytic amount of sulphuric acid at 80 8C. The chalcone intermediates 4a– d were obtained by treating aldehydes 2a–c with various substituted acetophenones under alkali media, and later they were reduced to corresponding hydroxyl derivatives 5a–d using sodium borohydride at room temperature. In another series, oxazolones (6a–f) were synthesized in good yield by treating substituted/unsubstituted hippuric acids with aldehydes 2a–c, in the presence of sodium acetate and acetic anhydride under reflux
1001-8417/$ – see front matter ß 2013 Airody Vasudeva Adhikari. Published by Elsevier B.V. on behalf of Chinese Chemical Society. All rights reserved. http://dx.doi.org/10.1016/j.cclet.2013.05.030
Please cite this article in press as: S. Ulloora, et al., Synthesis and antiepileptic studies of new imidazo[1,2-a]pyridine derivatives, Chin. Chem. Lett. (2013), http://dx.doi.org/10.1016/j.cclet.2013.05.030
G Model
CCLET-2620; No. of Pages 4 S. Ulloora et al. / Chinese Chemical Letters xxx (2013) xxx–xxx
2
N
N
3a: R1= F; R 2= Tolyl
R2N O
N
R2NH2, Ethanol
Br N
Ethanol
+
DMF-POCl 3, 0-5 oC
N
o
NH2
3b: R1= H; R 2= Tolyl 3c: R1= F; R 2= Benzothiazol-2-yl 3d: R1= H; R 2= Benzothiazol -2-yl
H+, 24 h
N
80 C, 6 h
R1 R1
1a-c
R1
3a-d
O
o
80 C, 6 h
N
R1= H, F, OMe
R3COCH3, 50 oC NaOH, 4 h
R1
2a-c
N
N
O R3
R4PhCONHCH 2CO 2H NCCH2CO2Et, Piperidine Ethanol, r.t.
NH2 N
Ac2O, NaOAc, 4 h R4
Ethanol
N
N
N NC
N
N
N
O
N
HN 8a-c
R1
N
4b, 5b: R1= F; R 3= Thiophen -2-yl 4c, 5c: R1= H; R 3= Thiophen -2-yl 4d, 5d: R1= OMe; R 3= 4-Chlorophenyl
O
EtOOC
O
R1
NaBH 4, r.t. 4a, 5a: R1= F; R 3= 4-Chlorophenyl Ethanol
N2H4.H2O
N
4a-d
6a-f
7a-c
7a, 8a: R1= H; 7b, 8b : R1= F; 7c, 8c: R1= OMe
R1
R1 6d: R1= H; R 4= Me 6a: R1, R4= H 6e: R1= F; R 4= Me 6b: R1= F; R 4= H 6c: R1= OMe; R 4= H 6f: R1= OMe; R 4= Me
HO R3 5a-d
R1
Scheme 1. Synthetic routes for target compounds.
condition. Further, the aldehydes 2a–c were coupled with ethyl cyanoacetate in the presence of piperidine to obtain 7a–c that later successfully cyclized to pyrazolone derivatives 8a–c by reacting with hydrazine hydrate. The detailed procedures followed for the synthesis of these new derivatives and their characterization data were given in Supporting information. The target compounds were screened for their in vivo anticonvulsant activity following maximal electroshock seizure
(MES) [13] and subcutaneous pentylene tetrazole (scPTZ) [14] methods, while their toxicity study was performed by the Rotarod technique [15]. Three different test doses, such as 20 mg/kg, 40 mg/kg and 100 mg/kg were used for these studies. These animal studies were performed in accordance with the ethical standards on animal experiments. The screening results are tabulated in Table 1. The procedures followed for these studies are discussed in the Supporting information.
Table 1 Anticonvulsant screening results of target compounds. Sample
3a 3b 3c 3d 5a 5b 5c 5d 6a 6b 6c 6d 6e 6f 7a 7b 7c 8a 8b 8c Phenytoin Diazepam
MESa
scPTZa
Toxicity resultsa
0.5
4.0
0.5
4.0
0.5
4.0
– – – – – – – – – – – – – 40 – – – – – – 20 x
– – 100 – – – – – – – – – – – – – – – – – 20 x
40 – 40 100 – 100 – – – 20 40 – 20 20 – – – – 40 100 x 20
– – 100 – – – – 100 – 100 100 – 40 40 – – 100 – 100 – x 20
– x – – x – – x x – – x – – x x – x – x 100 –
– x 100 – x – – x x – – x – – x x – x 100 x 100 –
a Doses of 20 mg/kg, 40 mg/kg, 100 mg/kg of the compounds were administered and the protection as well as toxicity were measured after 0.5 h and 4.0 h. The figures indicate the minimal concentration of sample required to cause either protection or toxicity in more than 50% of mice. The dash (–) indicates the absence of activity/toxicity, while (x) denotes not tested.
Please cite this article in press as: S. Ulloora, et al., Synthesis and antiepileptic studies of new imidazo[1,2-a]pyridine derivatives, Chin. Chem. Lett. (2013), http://dx.doi.org/10.1016/j.cclet.2013.05.030
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CCLET-2620; No. of Pages 4 S. Ulloora et al. / Chinese Chemical Letters xxx (2013) xxx–xxx
3. Results and discussion 3.1. Chemistry
3
showed activity at a dose of 40 mg/kg, while compounds 3d, 5b, 5d, 7c and 8c exhibited anticonvulsant activity at a relatively high test dose of 100 mg/kg. Based on general observations on structure–activity relationship, it appears that the presence of an electron donating substituent at the position-4 of the phenyl ring attached at C-2 of the imidazo[1,2-a]pyridine nucleus results in enhanced anticonvulsant activity. In particular, presence of fluoro (as in 3a, 3c, 5b, 6b, 6e and 8b) and methoxy (as in 6c, 6f, 7c and 8c) groups led to significant activity when compared with those of unsubstituted phenyl derivatives. Among various imidazo[1,2a]pyridine derivatives, oxazolone analogues emerged as lead compounds. Particularly, compounds 6e and 6f, possessing a tolyl substituent on the oxazolone pharmacophore, displayed enhanced activity when compared with those possessing a phenyl substituent (as in 6b and 6c). Thus, as electron donating strength of substituents present on the oxazolone moiety increases, their anticonvulsant activity also increases. Further, the pyrazolone derivatives 8a–c were found to be more active than their precursors (7a–c). This may be due to their capacity to involve in hydrogen bonding interactions with cellular receptors. Furthermore, the neurotoxicity screening results indicated that most of the tested compounds are non-toxic at tested doses 20 mg/kg, 40 mg/kg and 100 mg/kg. Only compounds 3c and 8b showed toxic character at a high dose of 100 mg/kg.
The structures of the new compounds were confirmed by FTIR, H NMR, 13C NMR, and mass spectrometry followed by elemental analysis studies. Conversion of aldehyde to Schiff base was confirmed by the FTIR spectrum of 3a, wherein the carbonyl peak due to aldehyde disappeared, while a new peak corresponding to C5 5N stretching appeared at 1607 cm 1. In its 1H NMR spectrum, a singlet peak at d 10.07 due to CH5 5N proton appeared, which further confirms the conversion. A peak at 1648 cm 1 was observed in the FTIR spectrum of chalcone 4a, which is due to stretching vibration of the conjugated carbonyl group. Upon reduction, this carbonyl peak has disappeared in FTIR spectrum of 5a, indicating the conversion of C5 5O to CHOH. Later, this reduction was further confirmed by its 1H NMR spectrum, where a new peak at d 2.02 has appeared due to a hydroxyl group. Also, three double doublets were observed at d 6.76, 6.14, 5.38, due to CH5 5CH and allylic protons. This clearly shows that only the carbonyl group had undergone reduction while the double bond was intact. FTIR spectrum of oxazolone derivative 6a showed a carbonyl peak at 1777 cm 1, indicating the presence of a cyclic lactone ring. Also, appearance of another peak at 1640 cm 1 due to C5 5N stretching of oxazolone further supported the proposed structure. Additional evidence was obtained by its 13C NMR spectrum where two characteristic peaks at d 168.5 and 165.4 due to carbonyl and C5 5N carbons of oxazolone moiety, respectively, were observed. Finally, conversion of aldehydes 2a–c into ester derivatives 7a– c was confirmed by FTIR spectrum of 7a, wherein characteristic peaks due to nitrile and carboxylic esters were observed at 2218 and 1710 cm 1, respectively. Its 1H NMR spectrum showed a singlet at d 8.31 due to vinylic CH proton, a quartet and a triplet at d 4.08 and 1.08 due to an ethyl carboxylate group. Similarly, the cyclization of ester analogues 7a–c to pyrazolones 8a–c was confirmed by their FTIR and 1H NMR spectral studies. The peaks due to the nitrile and ester groups of 7a disappeared in the FTIR spectrum of 8a, while new peaks at 3303 and 3176 cm 1 have appeared due to amine and amide NH groups, respectively. The peak due to cyclic conjugated amide carbonyl appeared at a relatively lower stretching frequency (1607 cm 1), which is due to conjugation involved with the cyclic carbamide group and the exocyclic double bond. Its 1H NMR spectrum displayed two singlets at d 9.89 and 2.84 due to NH and NH2 groups, respectively, confirming the conversion. Similarly, all target compounds were also well-characterized by FTIR, 1H NMR, 13C NMR and mass spectral studies.
The authors are thankful to National Institute of Technology Karnataka, India for financial support and laboratory facility, Indian Institute of Science, Bangalore for NMR facilities and SAIF Punjab for mass spectral facilities.
3.2. Anticonvulsant study
Appendix A. Supplementary data
1
The results of the MES screening indicated that compound 3c possessing benzothiazole ring is active at 100 mg/kg after 4 h of sample injection, while compound 6f containing oxazolone ring displays activity at a smaller dose of 40 mg/kg. Interestingly, new compounds displayed significant activity in the scPTZ method when compared to their activity in the MES method. This clearly indicates their ability to elevate the seizure threshold. Further, compounds 3a, 3c, 3d, 5b, 5d, 6b, 6c, 6e, 6f, 7c, 8b and 8c showed significant antiepileptic activity in the scPTZ method. Particularly, compounds 6b, 6e and 6f containing oxazolone moiety displayed very good activity at a dose of 20 mg/kg which is comparable to that of standard drug diazepam by preventing chemically induced seizures. Further, they were active in both the intervals (0.5 h and 4 h), indicating their rapid onset and long duration of anticonvulsant action. On the other hand, compounds 3a, 3c, 6c, and 8b
4. Conclusion New imidazo[1,2-a]pyridine derivatives carrying active pharmacophores at position-3 were successfully designed, synthesized and characterized by various spectral methods. The target compounds were subjected to in vivo antiepileptic and toxicity studies following standard methods. New compounds exhibited better activity in the scPTZ method than the MES method, indicating their ability to elevate seizure threshold effectively. Compounds 6e and 6f possessing tolyl substituted oxazolone moiety emerged as lead compounds by displaying prominent anticonvulsant activity in the scPTZ method at a dose of 20 mg/kg. Most of the tested compounds were found to be non-toxic at all tested doses. Acknowledgments
Supplementary data associated with this article can be found, in the online version, at http://dx.doi.org/10.1016/j.cclet.2013.05.030. References [1] A. Karakurt, M. Ozalp, S. Isik, J.P. Stables, S. Dalkara, Synthesis, anticonvulsant and antimicrobial activities of some new 2-acetylnaphthalene derivatives, Bioorg. Med. Chem. 18 (2010) 2902–2911. [2] B.M. Kenda, A.C. Matagne, P.E. Talaga, et al., Discovery of 4-substituted pyrrolidone butanamides as new agents with significant antiepileptic activity, J. Med. Chem. 47 (2004) 530–549. [3] M.C. Picot, M. Baldy-Moulinier, J.P. Daurs, P. Dujols, A. Crespel, The prevalence of epilepsy and pharmacoresistant epilepsy in adults: a population based study in a western European country, Epilepsia 49 (2008) 1230–1238. [4] M. Naithani, S. Chopra, B.L. Somani, R.K. Singh, Studies on adverse metabolic effects of antiepileptics and their correlation with blood components, Curr. Neurobiol. 01 (2010) 117–120.
Please cite this article in press as: S. Ulloora, et al., Synthesis and antiepileptic studies of new imidazo[1,2-a]pyridine derivatives, Chin. Chem. Lett. (2013), http://dx.doi.org/10.1016/j.cclet.2013.05.030
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[5] W. Haefely, J.R. Martin, P. Schoch, Novel anxiolytics that act as partial agonists at benzodiazepine receptors, Trends Pharmacol. Sci. 11 (1990) 452–456. [6] J. Vlainic, D. Pericic, Zolpidem is a potent anticonvulsant in adult and aged mice, Brain Res. 1310 (2010) 181–188. [7] C. Hamdouchi, J. de Blas, M. del Prado, et al., 2-Amino-3-substituted-6-[(E)-1phenyl-2-N-methyl carbamoyl)vinyl]imidazo[1 2-a]pyridines as a novel class of inhibitors of human rhinovirus: stereospecific synthesis and antiviral activity, J. Med. Chem. 42 (1999) 50–59. [8] N.D. Farkas, C. Langley, A.L. Rousseau, et al., 6-Substituted imidazo[1 2-a]pyridines: synthesis and biological activity against colon cancer cell lines HT-29 and Caco-2, Eur. J. Med. Chem. 46 (2011) 4573–4583. [9] N. Denora, V. Laquintana, M.G. Pisu, et al., 2-Phenyl imidazo[1 2-a]pyridine compounds containing hydrophilic groups as potent and selective ligands for peripheral benzodiazepine receptors: synthesis, binding affinity and electrophysiological studies, J. Med. Chem. 51 (2008) 6876–6888.
[10] A.S. El-Azab, K.E.B. ElTahir, Design and synthesis of novel 7-aminoquinazoline derivatives: antitumor and anticonvulsant activities, Bioorg. Med. Chem. Lett. 22 (2012) 1879–1885. [11] H. Joshi, P. Upadhyay, D. Karia, A.J. Baxi, Synthesis of some novel imidazolinones as potent anticonvulsant agents, Eur. J. Med. Chem. 38 (2003) 837–840. [12] S. Ulloora, R. Shabaraya, A.V. Adhikari, New imidazo[1 2-a]pyridines carrying active pharmacophores: synthesis and anticonvulsant studies, Bioorg. Med. Chem. Lett. 23 (2013) 1502–1506. [13] R.K. Krall, J.K. Penry, B.G. White, H.J. Kupferberg, E.A. Swinyard, Antiepileptic drug development. II: anticonvulsant drug screening, Epilepsia 19 (1978) 409–428. [14] C.R. Clark, M.J.M. Wells, R.T. Sansom, et al., Anticonvulsant activity of some 4aminobenzamides, J. Med. Chem. 27 (1984) 779–782. [15] N.W. Dunham, T.S. Miya, A note on a simple apparatus for detecting neurological deficit in rats and mice, J. Am. Pharm. Assoc. Sci. Ed. 46 (1957) 208–209.
Please cite this article in press as: S. Ulloora, et al., Synthesis and antiepileptic studies of new imidazo[1,2-a]pyridine derivatives, Chin. Chem. Lett. (2013), http://dx.doi.org/10.1016/j.cclet.2013.05.030
CCLET-2620; No. of Pages 4 Chinese Chemical Letters xxx (2013) xxx–xxx
Contents lists available at SciVerse ScienceDirect
Chinese Chemical Letters journal homepage: www.elsevier.com/locate/cclet
Original article
Synthesis and antiepileptic studies of new imidazo[1,2-a]pyridine derivatives Shrikanth Ulloora a, Airody Vasudeva Adhikari a,*, Ramakrishna Shabaraya b a b
Department of Chemistry, National Institute of Technology Karnataka, Surathkal, Mangalore 575025, Karnataka, India Srinivas College of Pharmacy, Valachil, Mangalore, Karnataka, India
A R T I C L E I N F O
A B S T R A C T
Article history: Received 7 March 2013 Received in revised form 6 May 2013 Accepted 17 May 2013 Available online xxx
In this manuscript, we report the synthesis of newly designed imidazo[1,2-a]pyridines carrying active pharmacophores as potential anticonvulsant agents. Newly synthesized target compounds were characterized by FTIR, 1H NMR, 13C NMR, and mass spectroscopy followed by elemental analysis studies. Preliminary anticonvulsant screening study of target compounds was carried out following MES and scPTZ methods. Compounds 6e and 6f, possessing electron rich aryl substituent at position-2 and tolyl substituted oxazolone moiety at the position-3 of imidazo[1,2-a]pyridine ring, exhibited activity comparable to standard drug diazepam and emerged as lead compounds. New compounds displayed enhanced activity in scPTZ method, indicating their ability to raise the seizure threshold. Their neurotoxicity study by Rotarod test showed that they are nontoxic at all tested doses. ß 2013 Airody Vasudeva Adhikari. Published by Elsevier B.V. on behalf of Chinese Chemical Society. All rights reserved.
Keywords: Anticonvulsant study Imidazo[1,2-a]pyridine In vivo study Neurotoxicity
1. Introduction Epilepsy is a major neurological disease, affecting about 50 million people worldwide [1]. The cellular mechanism of human epilepsy is still uncertain and hence present drug therapy only treats epileptic symptoms, rather than curing it [2]. Now there are more than 40 different anti-epileptic drugs (AEDs) in clinical use, but still about 30% of patients continue to experience uncontrolled seizures, and they are pharmaco-resistant to the available therapy [3]. Further, present medication for seizures requires continuous medication for a long period, which is in turn associated with many adverse side effects, such as nausea, ataxia, drowsiness, gastrointestinal disturbance, hyperplasia, anaemia etc. [4]. As a result, design and development of new, efficient antiepileptic agents has become an active area of research in medicinal chemistry. Imidazo[1,2-a]pyridines are an important class of non-benzodiazepines, exhibiting very good CNS activities and showing fewer adverse effects compared to classical benzodiazepines [5]. Some of the imidazo[1,2-a]pyridine derivatives, such as Zolpidem, Alpidem, Saripidem and DS-1, are well-known CNS drugs, exhibiting antiepileptic activity at lesser doses [6]. The pharmacological profiles of imidazo[1,2-a]pyridines are mainly dependent on the nature of the substituents at their position-2 and position-3 [7]. The imidazo[1,2-a]pyridines carrying aryl groups at the position-2 are reported as highly CNS active
* Corresponding author. E-mail address: [email protected] (A.V. Adhikari).
scaffolds [8]. Further, it is also revealed that, such scaffolds possessing a hydrophobic unit at the position-8 along with a halogen substituent on the aryl ring can bind selectively to benzodiazepine receptors [9]. As such, this work presents new imidazo[1,2-a]pyridines carrying a halo substituted aryl ring at the position-2 along with a hydrophobic methyl group at the position8. Additionally, active pharmacophoric groups like oxazolones, pyrazolones, imines and hydroxyl derivatives have been incorporated at the position-3 owing to their established prominent anticonvulsant activities [10,11]. Accordingly, four new series of imidazo[1,2-a]pyridines carrying imines (3a–d), enol derivatives (5a–d), oxazolones (6a–f), carboxylate intermediates (7a–c) and pyrazolones (8a–c) have been synthesized following appropriate synthetic routes as shown in Scheme 1. 2. Experimental The synthesis and the characterization details of intermediates (1a–c and 2a–c) were reported in our previous publication [12]. The imines 3a–d were obtained by condensing aldehydes 2a–c with appropriate aromatic amines in the presence of a catalytic amount of sulphuric acid at 80 8C. The chalcone intermediates 4a– d were obtained by treating aldehydes 2a–c with various substituted acetophenones under alkali media, and later they were reduced to corresponding hydroxyl derivatives 5a–d using sodium borohydride at room temperature. In another series, oxazolones (6a–f) were synthesized in good yield by treating substituted/unsubstituted hippuric acids with aldehydes 2a–c, in the presence of sodium acetate and acetic anhydride under reflux
1001-8417/$ – see front matter ß 2013 Airody Vasudeva Adhikari. Published by Elsevier B.V. on behalf of Chinese Chemical Society. All rights reserved. http://dx.doi.org/10.1016/j.cclet.2013.05.030
Please cite this article in press as: S. Ulloora, et al., Synthesis and antiepileptic studies of new imidazo[1,2-a]pyridine derivatives, Chin. Chem. Lett. (2013), http://dx.doi.org/10.1016/j.cclet.2013.05.030
G Model
CCLET-2620; No. of Pages 4 S. Ulloora et al. / Chinese Chemical Letters xxx (2013) xxx–xxx
2
N
N
3a: R1= F; R 2= Tolyl
R2N O
N
R2NH2, Ethanol
Br N
Ethanol
+
DMF-POCl 3, 0-5 oC
N
o
NH2
3b: R1= H; R 2= Tolyl 3c: R1= F; R 2= Benzothiazol-2-yl 3d: R1= H; R 2= Benzothiazol -2-yl
H+, 24 h
N
80 C, 6 h
R1 R1
1a-c
R1
3a-d
O
o
80 C, 6 h
N
R1= H, F, OMe
R3COCH3, 50 oC NaOH, 4 h
R1
2a-c
N
N
O R3
R4PhCONHCH 2CO 2H NCCH2CO2Et, Piperidine Ethanol, r.t.
NH2 N
Ac2O, NaOAc, 4 h R4
Ethanol
N
N
N NC
N
N
N
O
N
HN 8a-c
R1
N
4b, 5b: R1= F; R 3= Thiophen -2-yl 4c, 5c: R1= H; R 3= Thiophen -2-yl 4d, 5d: R1= OMe; R 3= 4-Chlorophenyl
O
EtOOC
O
R1
NaBH 4, r.t. 4a, 5a: R1= F; R 3= 4-Chlorophenyl Ethanol
N2H4.H2O
N
4a-d
6a-f
7a-c
7a, 8a: R1= H; 7b, 8b : R1= F; 7c, 8c: R1= OMe
R1
R1 6d: R1= H; R 4= Me 6a: R1, R4= H 6e: R1= F; R 4= Me 6b: R1= F; R 4= H 6c: R1= OMe; R 4= H 6f: R1= OMe; R 4= Me
HO R3 5a-d
R1
Scheme 1. Synthetic routes for target compounds.
condition. Further, the aldehydes 2a–c were coupled with ethyl cyanoacetate in the presence of piperidine to obtain 7a–c that later successfully cyclized to pyrazolone derivatives 8a–c by reacting with hydrazine hydrate. The detailed procedures followed for the synthesis of these new derivatives and their characterization data were given in Supporting information. The target compounds were screened for their in vivo anticonvulsant activity following maximal electroshock seizure
(MES) [13] and subcutaneous pentylene tetrazole (scPTZ) [14] methods, while their toxicity study was performed by the Rotarod technique [15]. Three different test doses, such as 20 mg/kg, 40 mg/kg and 100 mg/kg were used for these studies. These animal studies were performed in accordance with the ethical standards on animal experiments. The screening results are tabulated in Table 1. The procedures followed for these studies are discussed in the Supporting information.
Table 1 Anticonvulsant screening results of target compounds. Sample
3a 3b 3c 3d 5a 5b 5c 5d 6a 6b 6c 6d 6e 6f 7a 7b 7c 8a 8b 8c Phenytoin Diazepam
MESa
scPTZa
Toxicity resultsa
0.5
4.0
0.5
4.0
0.5
4.0
– – – – – – – – – – – – – 40 – – – – – – 20 x
– – 100 – – – – – – – – – – – – – – – – – 20 x
40 – 40 100 – 100 – – – 20 40 – 20 20 – – – – 40 100 x 20
– – 100 – – – – 100 – 100 100 – 40 40 – – 100 – 100 – x 20
– x – – x – – x x – – x – – x x – x – x 100 –
– x 100 – x – – x x – – x – – x x – x 100 x 100 –
a Doses of 20 mg/kg, 40 mg/kg, 100 mg/kg of the compounds were administered and the protection as well as toxicity were measured after 0.5 h and 4.0 h. The figures indicate the minimal concentration of sample required to cause either protection or toxicity in more than 50% of mice. The dash (–) indicates the absence of activity/toxicity, while (x) denotes not tested.
Please cite this article in press as: S. Ulloora, et al., Synthesis and antiepileptic studies of new imidazo[1,2-a]pyridine derivatives, Chin. Chem. Lett. (2013), http://dx.doi.org/10.1016/j.cclet.2013.05.030
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CCLET-2620; No. of Pages 4 S. Ulloora et al. / Chinese Chemical Letters xxx (2013) xxx–xxx
3. Results and discussion 3.1. Chemistry
3
showed activity at a dose of 40 mg/kg, while compounds 3d, 5b, 5d, 7c and 8c exhibited anticonvulsant activity at a relatively high test dose of 100 mg/kg. Based on general observations on structure–activity relationship, it appears that the presence of an electron donating substituent at the position-4 of the phenyl ring attached at C-2 of the imidazo[1,2-a]pyridine nucleus results in enhanced anticonvulsant activity. In particular, presence of fluoro (as in 3a, 3c, 5b, 6b, 6e and 8b) and methoxy (as in 6c, 6f, 7c and 8c) groups led to significant activity when compared with those of unsubstituted phenyl derivatives. Among various imidazo[1,2a]pyridine derivatives, oxazolone analogues emerged as lead compounds. Particularly, compounds 6e and 6f, possessing a tolyl substituent on the oxazolone pharmacophore, displayed enhanced activity when compared with those possessing a phenyl substituent (as in 6b and 6c). Thus, as electron donating strength of substituents present on the oxazolone moiety increases, their anticonvulsant activity also increases. Further, the pyrazolone derivatives 8a–c were found to be more active than their precursors (7a–c). This may be due to their capacity to involve in hydrogen bonding interactions with cellular receptors. Furthermore, the neurotoxicity screening results indicated that most of the tested compounds are non-toxic at tested doses 20 mg/kg, 40 mg/kg and 100 mg/kg. Only compounds 3c and 8b showed toxic character at a high dose of 100 mg/kg.
The structures of the new compounds were confirmed by FTIR, H NMR, 13C NMR, and mass spectrometry followed by elemental analysis studies. Conversion of aldehyde to Schiff base was confirmed by the FTIR spectrum of 3a, wherein the carbonyl peak due to aldehyde disappeared, while a new peak corresponding to C5 5N stretching appeared at 1607 cm 1. In its 1H NMR spectrum, a singlet peak at d 10.07 due to CH5 5N proton appeared, which further confirms the conversion. A peak at 1648 cm 1 was observed in the FTIR spectrum of chalcone 4a, which is due to stretching vibration of the conjugated carbonyl group. Upon reduction, this carbonyl peak has disappeared in FTIR spectrum of 5a, indicating the conversion of C5 5O to CHOH. Later, this reduction was further confirmed by its 1H NMR spectrum, where a new peak at d 2.02 has appeared due to a hydroxyl group. Also, three double doublets were observed at d 6.76, 6.14, 5.38, due to CH5 5CH and allylic protons. This clearly shows that only the carbonyl group had undergone reduction while the double bond was intact. FTIR spectrum of oxazolone derivative 6a showed a carbonyl peak at 1777 cm 1, indicating the presence of a cyclic lactone ring. Also, appearance of another peak at 1640 cm 1 due to C5 5N stretching of oxazolone further supported the proposed structure. Additional evidence was obtained by its 13C NMR spectrum where two characteristic peaks at d 168.5 and 165.4 due to carbonyl and C5 5N carbons of oxazolone moiety, respectively, were observed. Finally, conversion of aldehydes 2a–c into ester derivatives 7a– c was confirmed by FTIR spectrum of 7a, wherein characteristic peaks due to nitrile and carboxylic esters were observed at 2218 and 1710 cm 1, respectively. Its 1H NMR spectrum showed a singlet at d 8.31 due to vinylic CH proton, a quartet and a triplet at d 4.08 and 1.08 due to an ethyl carboxylate group. Similarly, the cyclization of ester analogues 7a–c to pyrazolones 8a–c was confirmed by their FTIR and 1H NMR spectral studies. The peaks due to the nitrile and ester groups of 7a disappeared in the FTIR spectrum of 8a, while new peaks at 3303 and 3176 cm 1 have appeared due to amine and amide NH groups, respectively. The peak due to cyclic conjugated amide carbonyl appeared at a relatively lower stretching frequency (1607 cm 1), which is due to conjugation involved with the cyclic carbamide group and the exocyclic double bond. Its 1H NMR spectrum displayed two singlets at d 9.89 and 2.84 due to NH and NH2 groups, respectively, confirming the conversion. Similarly, all target compounds were also well-characterized by FTIR, 1H NMR, 13C NMR and mass spectral studies.
The authors are thankful to National Institute of Technology Karnataka, India for financial support and laboratory facility, Indian Institute of Science, Bangalore for NMR facilities and SAIF Punjab for mass spectral facilities.
3.2. Anticonvulsant study
Appendix A. Supplementary data
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The results of the MES screening indicated that compound 3c possessing benzothiazole ring is active at 100 mg/kg after 4 h of sample injection, while compound 6f containing oxazolone ring displays activity at a smaller dose of 40 mg/kg. Interestingly, new compounds displayed significant activity in the scPTZ method when compared to their activity in the MES method. This clearly indicates their ability to elevate the seizure threshold. Further, compounds 3a, 3c, 3d, 5b, 5d, 6b, 6c, 6e, 6f, 7c, 8b and 8c showed significant antiepileptic activity in the scPTZ method. Particularly, compounds 6b, 6e and 6f containing oxazolone moiety displayed very good activity at a dose of 20 mg/kg which is comparable to that of standard drug diazepam by preventing chemically induced seizures. Further, they were active in both the intervals (0.5 h and 4 h), indicating their rapid onset and long duration of anticonvulsant action. On the other hand, compounds 3a, 3c, 6c, and 8b
4. Conclusion New imidazo[1,2-a]pyridine derivatives carrying active pharmacophores at position-3 were successfully designed, synthesized and characterized by various spectral methods. The target compounds were subjected to in vivo antiepileptic and toxicity studies following standard methods. New compounds exhibited better activity in the scPTZ method than the MES method, indicating their ability to elevate seizure threshold effectively. Compounds 6e and 6f possessing tolyl substituted oxazolone moiety emerged as lead compounds by displaying prominent anticonvulsant activity in the scPTZ method at a dose of 20 mg/kg. Most of the tested compounds were found to be non-toxic at all tested doses. Acknowledgments
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